The adaptational response of the heart to an imposed load is complex and engages multiple intra and extracellular signaling processes, the net effect of which is to alter the cardiac phenotype. During the first cycle of funding, we have recapitulated cardiac (mal)adaptation by expressing single regulatory kinases (PKCe and PKCP) in the murine heart at physiologically relevant levels. Analysis of these gain-of-function mice has revealed separable and distinct patterns of response that reflect different biochemical and molecular mechanisms. Expression of PKCp causes concentric hypertrophy with impaired relaxation associated with abnormalities in calcium handling and is largely independent of changes in myofibrillar protein composition. In contrast, expression of PKCs results in the gradual onset of a dilated, non-hypertrophic myopathy with depressed contractility associated with changes in contractile protein composition but without changes in calcium availability. These findings suggest several hypotheses that will be explored during the next period of funding: First, the progression to ventricular dilatation reflects a shifted relationship among phosphoproteins within the myofilament (in particular, troponin 1 and the regulatory myosin light chain) that change overall calcium sensitivity and cross-bridge cycling properties. Second, the progression from one stage of adaptation to the next (and ultimately to heart failure) is determined both by the signaling pathways that are engaged and also by the cellular context and the changing combinatorial targets upon which these signaling pathways are acting. Third, correction of this sarcomeric protein imbalance has the potential to normalize the contractile properties and prevent and/or reverse the progression of dilated cardiomyopathy. To explore these hypotheses we will first characterize sarcomeric abnormalities that herald compensatory and decompensatory moments in the adaptational response to PKC activation. A targeted sub-proteomic strategy will be used coupled with biochemical analysis to establish relevant biochemical and molecular changes that are associated with defined mechanical measurements. Subsequently we will specifically repair key features of sarcomeric maladaptation and establish if this reverses or prevents the development of the pathologic phenotype seen in PKCe mice. Multiple strategies, including myofilament protein exchange, in situ protein transfer, and transgenic crosses will be employed. Finally, we will generalize these biochemical findings to human disease by examining biopsy samples taken from carefully defined patient populations during open heart surgery.